Abstract：
OBJECTIVE: Mechanisms and clinical impact of portal microthrombosis featuring severe COVID-19 are unknown. Intrapulmonary vascular dilation (IPVD)-related hypoxia has been described in severe liver diseases. We hypothesized that portal microthrombosis is associated with IPVD and fatal respiratory failure in COVID-19.
METHODS: Ninety-three patients who died from COVID-19, were analysed for portal microvascular damage (histology), IPVD (histology and chest-computed tomography, CT), and hypoxemia (arterial blood gas). Seventeen patients who died from COVID-19-unrelated pneumonia served as controls. Vascular lesions and microthrombi were phenotyped for endothelial (vWF) and pericyte (αSMA/PDGFR-β) markers, tissue factor (TF), viral spike-protein and nucleoprotein (SP, NP), fibrinogen, platelets (CD41a). Viral particles in vascular cells were assessed by transmission electron microscopy (TEM). Cultured pericytes were infected with SARS-CoV-2 to measure TF expression and tubulisation of human pulmonary microvascular endothelial cells (HPMEC) was assessed upon vWF treatment.
RESULTS: IPVD was present in 16/66 COVID-19 patients with both liver and lung histology, with a younger age (62 vs 78yo), longer illness (25 vs 14 days), worsening hypoxemia (PaO2/FiO2 from 209 to 89), and more ventilatory support (63 vs 22%) compared to COVID-19/Non-IPVD. IPVD, absent in controls, were confirmed by chest-CT. COVID-19/IPVD liver histology showed portal microthrombosis in >82.5% of portal areas, with a thicker wall of αSMA/PDGFR-β+/ SP+/NP+ pericytes compared with COVID-19/Non-IPVD. Thrombosed portal venules correlated with αSMA+ area, whereas infected SP+/NP+ pericytes expressed TF. SARS-CoV-2 viral particles were observed in portal pericytes. In-vitro SARS-CoV-2 infection of pericytes up-regulated TF and induced endothelial cells to overexpress vWF, which expanded HPMEC tubules.
CONCLUSIONS: SARS-CoV-2 infection of liver pericytes elicits a local procoagulant response associated with extensive portal microthrombosis, IPVD and worsening respiratory failure in fatal COVID-19.
UNASSIGNED: Vascular involvement of the liver represents a serious complication of COVID-19 infection that must be considered in the work-up of patients with long-lasting and progressively worsening respiratory failure, as it may associate with the development of intrapulmonary vascular dilations. This clinical picture is associated with a pro-coagulant phenotype of portal venule pericytes, which is induced by SARS-CoV-2 infection of pericytes. Both observations provide a model that may apply, at least in part, to other vascular disorders of the liver, featuring obliterative portal venopathy, similarly characterized at the clinical level by development of hypoxemia and at the histological level, by phlebosclerosis and reduced caliber of the portal vein branches in the absence of cirrhosis. Moreover, our findings bring light to an as yet overlooked player of thrombosis pathophysiology, i.e. pericytes, which may provide novel therapeutic tools to halt prothrombotic mechanisms.
METHODS: Ninety-three patients who died from COVID-19, were analysed for portal microvascular damage (histology), IPVD (histology and chest-computed tomography, CT), and hypoxemia (arterial blood gas). Seventeen patients who died from COVID-19-unrelated pneumonia served as controls. Vascular lesions and microthrombi were phenotyped for endothelial (vWF) and pericyte (αSMA/PDGFR-β) markers, tissue factor (TF), viral spike-protein and nucleoprotein (SP, NP), fibrinogen, platelets (CD41a). Viral particles in vascular cells were assessed by transmission electron microscopy (TEM). Cultured pericytes were infected with SARS-CoV-2 to measure TF expression and tubulisation of human pulmonary microvascular endothelial cells (HPMEC) was assessed upon vWF treatment.
RESULTS: IPVD was present in 16/66 COVID-19 patients with both liver and lung histology, with a younger age (62 vs 78yo), longer illness (25 vs 14 days), worsening hypoxemia (PaO2/FiO2 from 209 to 89), and more ventilatory support (63 vs 22%) compared to COVID-19/Non-IPVD. IPVD, absent in controls, were confirmed by chest-CT. COVID-19/IPVD liver histology showed portal microthrombosis in >82.5% of portal areas, with a thicker wall of αSMA/PDGFR-β+/ SP+/NP+ pericytes compared with COVID-19/Non-IPVD. Thrombosed portal venules correlated with αSMA+ area, whereas infected SP+/NP+ pericytes expressed TF. SARS-CoV-2 viral particles were observed in portal pericytes. In-vitro SARS-CoV-2 infection of pericytes up-regulated TF and induced endothelial cells to overexpress vWF, which expanded HPMEC tubules.
CONCLUSIONS: SARS-CoV-2 infection of liver pericytes elicits a local procoagulant response associated with extensive portal microthrombosis, IPVD and worsening respiratory failure in fatal COVID-19.
UNASSIGNED: Vascular involvement of the liver represents a serious complication of COVID-19 infection that must be considered in the work-up of patients with long-lasting and progressively worsening respiratory failure, as it may associate with the development of intrapulmonary vascular dilations. This clinical picture is associated with a pro-coagulant phenotype of portal venule pericytes, which is induced by SARS-CoV-2 infection of pericytes. Both observations provide a model that may apply, at least in part, to other vascular disorders of the liver, featuring obliterative portal venopathy, similarly characterized at the clinical level by development of hypoxemia and at the histological level, by phlebosclerosis and reduced caliber of the portal vein branches in the absence of cirrhosis. Moreover, our findings bring light to an as yet overlooked player of thrombosis pathophysiology, i.e. pericytes, which may provide novel therapeutic tools to halt prothrombotic mechanisms.
摘要：
目的：以严重COVID-19为特征的门脉微血栓形成的机制和临床影响未知。肺内血管扩张（IPVD）相关的缺氧已在严重的肝脏疾病中得到描述。我们假设门脉微血栓形成与COVID-19的IPVD和致命性呼吸衰竭有关。
方法：分析93例死于COVID-19的患者的门静脉微血管损伤（组织学），IPVD（组织学和胸部计算机断层扫描，CT),和低氧血症（动脉血气）。17例死于COVID-19无关肺炎的患者作为对照。血管病变和微血栓表型为内皮（vWF）和周细胞（αSMA/PDGFR-β）标记，组织因子（TF），病毒刺突蛋白和核蛋白(SP，NP),纤维蛋白原，血小板（CD41a）。通过透射电子显微镜(TEM)评估血管细胞中的病毒颗粒。用SARS-CoV-2感染培养的周细胞以测量TF表达，并在vWF治疗后评估人肺微血管内皮细胞（HPMEC）的雾化。
结果：16/66COVID-19患者肝和肺组织学均存在IPVD，年龄较小（62岁vs78岁），患病时间更长（25天vs14天），低氧血症恶化（PaO2/FiO2从209到89），与COVID-19/非IPVD相比，通气支持更多（63%vs22%）。IPVD,不在控件中，经胸部CT证实。COVID-19/IPVD肝组织学显示门脉微血栓形成>82.5%，与COVID-19/非IPVD相比，αSMA/PDGFR-β+/SP+/NP+周细胞壁较厚。血栓门静脉小静脉与αSMA+面积相关，而感染的SP+/NP+周细胞表达TF。在门静脉周细胞中观察到SARS-CoV-2病毒颗粒。体外SARS-CoV-2感染周细胞上调TF,诱导内皮细胞过表达vWF,扩大HPMEC小管。
结论：肝脏周细胞的SARS-CoV-2感染引起与广泛的门静脉微血栓形成相关的局部促凝血反应，致命COVID-19的IPVD和呼吸衰竭恶化。
肝脏血管受累是COVID-19感染的严重并发症，在治疗长期且逐渐恶化的呼吸衰竭患者时必须考虑到这一点，因为它可能与肺内血管扩张的发展有关。此临床表现与门静脉小静脉周细胞的促凝表型有关，这是由SARS-CoV-2感染周细胞引起的。这两种观察都提供了一个可能适用的模型，至少在某种程度上,肝脏的其他血管疾病，以闭塞性门脉静脉病为特征，在临床水平和组织学水平上类似的特点是低氧血症的发展，在没有肝硬化的情况下，静脉硬化和门静脉分支的口径减小。此外，我们的发现为尚未被忽视的血栓形成病理生理学参与者带来了启示，即周皮细胞,这可能提供新的治疗工具来阻止血栓形成机制。
方法：分析93例死于COVID-19的患者的门静脉微血管损伤（组织学），IPVD（组织学和胸部计算机断层扫描，CT),和低氧血症（动脉血气）。17例死于COVID-19无关肺炎的患者作为对照。血管病变和微血栓表型为内皮（vWF）和周细胞（αSMA/PDGFR-β）标记，组织因子（TF），病毒刺突蛋白和核蛋白(SP，NP),纤维蛋白原，血小板（CD41a）。通过透射电子显微镜(TEM)评估血管细胞中的病毒颗粒。用SARS-CoV-2感染培养的周细胞以测量TF表达，并在vWF治疗后评估人肺微血管内皮细胞（HPMEC）的雾化。
结果：16/66COVID-19患者肝和肺组织学均存在IPVD，年龄较小（62岁vs78岁），患病时间更长（25天vs14天），低氧血症恶化（PaO2/FiO2从209到89），与COVID-19/非IPVD相比，通气支持更多（63%vs22%）。IPVD,不在控件中，经胸部CT证实。COVID-19/IPVD肝组织学显示门脉微血栓形成>82.5%，与COVID-19/非IPVD相比，αSMA/PDGFR-β+/SP+/NP+周细胞壁较厚。血栓门静脉小静脉与αSMA+面积相关，而感染的SP+/NP+周细胞表达TF。在门静脉周细胞中观察到SARS-CoV-2病毒颗粒。体外SARS-CoV-2感染周细胞上调TF,诱导内皮细胞过表达vWF,扩大HPMEC小管。
结论：肝脏周细胞的SARS-CoV-2感染引起与广泛的门静脉微血栓形成相关的局部促凝血反应，致命COVID-19的IPVD和呼吸衰竭恶化。
肝脏血管受累是COVID-19感染的严重并发症，在治疗长期且逐渐恶化的呼吸衰竭患者时必须考虑到这一点，因为它可能与肺内血管扩张的发展有关。此临床表现与门静脉小静脉周细胞的促凝表型有关，这是由SARS-CoV-2感染周细胞引起的。这两种观察都提供了一个可能适用的模型，至少在某种程度上,肝脏的其他血管疾病，以闭塞性门脉静脉病为特征，在临床水平和组织学水平上类似的特点是低氧血症的发展，在没有肝硬化的情况下，静脉硬化和门静脉分支的口径减小。此外，我们的发现为尚未被忽视的血栓形成病理生理学参与者带来了启示，即周皮细胞,这可能提供新的治疗工具来阻止血栓形成机制。
